1. From Gene to Protein

2. Gene Expression Process by which DNA directs the synthesis of a protein 2 stages transcription translation All organisms One gene  one protein

3. 1. Transcription of DNA Gene – Composed of DNA – 1000s of base pairs long – Sequence of G, A, T, C is a code A portion of the insulin gene ACATACATGTAGCAACGAAACCTGAACATGCGAACATTGACACAACAACAAUGUAGCCA

4. A gene is transcribed into mRNA – One DNA strand is a template – Follow base complementation using G, A, U, C Which strand is the template strand?

5.  This mRNA is referred to as a “transcript” Single stranded Uracil nucleotide (not thymine) ONLY gene of interest is transcribed mRNA will leave the nucleus for translation step

6. mRNA (transcript) leaves nucleus Translated to protein in cytoplasm What organelle translates mRNA to protein? How many amino acids? How many amino acids long is a protein? How could 10 identical protein molecules be translated from 1 mRNA? What happens to mRNA when the cell has finished with it? Can the cell transcribe more than one gene at a time?

7. 2. Translation of mRNA Nucleotides (G, A, U, C) translated to amino acids Ribosomes assemble amino acids to form polypeptide What is the difference between a polypeptide and a protein? What is the length difference between a chromosome and an mRNA transcript? Why do researchers use frog embryos to study translation?

8. Differences between prokaryotes and eukaryotes Proks – No nucleus – Translation can begin while transcription is still in progress

10. Euks – Nuclear envelope – Transcript is modified before leaving nucleus Primary transcript mRNA  mRNA – Translation in cytoplasm

12. Central Dogma DNA  mRNA  protein (F. Crick 1956)

13. The genetic code Triplet code 4 3 = – # combinations of 3 using G, A, T, C – Can code for all 20 aa Each triplet codes for a certain amino acid

14. Codon table (1960’s Nirenberg and Khorana) 3 stop codons 1 start codon Redundant Nearly universal

15. Which strand of DNA is the template? Identify the codons (triplets) Code is non-overlapping Compare the mRNA to the non-template strand How many codons are needed for a 20 amino acid long protein? How many nucleotides?

16. The reading frame AUG UGG GUU GGC UCA Amino acids? met –trp- phe-gly-ser

17. Transcription in more detail 1.Initiation of transcription A. RNA polymerase binds to template strand of DNA  upstream ----------------------------------------------downstream 

18. The promoter is upstream from the start site for transcription

19. B. Transcription factors (proteins) are required for RNA pol to bind

20. C. mRNA begins to be synthesized ( RNA pol does not need a primer)

21. 2. Elongation of transcript – RNA pol synthesizes 5’  3’ – More than one transcript made

22. 3. Termination RNA pol detaches

23. Translation summary

24. Translation in more detail  Cytoplasm The players: Transfer RNA tRNA Ribosomes mRNA Amino acids

25. 1. tRNA 2 o structure is a cloverleaf CONCEPT CHECK How many bases is a tRNA? How many bases is an anticodon? Where is the amino acid site with respect to the anticodon? Why does the tRNA look to be double stranded? What does the “t” mean in tRNA?

26. A charged tRNA carries an amino acid Contains an anticodon – Complementary to a codon Note the CCA

27. rRNA + protein Large and small subunits 2. Ribosome

28. 3 binding sites on ribosome P site holds tRNA A site for tRNA with next amino acid E site allows tRNA to leave Note: large and small subunits

29. Ribosomes adds each aa from tRNA to growing polypeptide chain up to 100,000s per cell Activity overview of protein synthesis

30. Stages of translation 1.Initiation Ribosome scans mRNA for AUG start tRNA brings met

32. 2. Elongation Amino acids added to chain Peptide bonds between amino acids

34. 3. Termination stop codon Polypeptide released

36. Polyribosomes One mRNA translated by many ribosomes

37. Wobble hypothesis How many sense codons? Stop codons? BUT……………only 45 tRNAs Some tRNAs recognize more than one codon

38. 3 rd position in mRNA codon “wobbles” Us and Cs can be matched with G in anticodon Rules of base pairing relaxed in the 3 rd position

39. After translation  ER  Golgi Post – translational modification